Initial sequencing and analysis of the human genome.
TL;DR: The results of an international collaboration to produce and make freely available a draft sequence of the human genome are reported and an initial analysis is presented, describing some of the insights that can be gleaned from the sequence.
Abstract: The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.
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TL;DR: The definition and use of family-specific, manually curated gathering thresholds are explained and some of the features of domains of unknown function (also known as DUFs) are discussed, which constitute a rapidly growing class of families within Pfam.
Abstract: Pfam is a widely used database of protein families and domains. This article describes a set of major updates that we have implemented in the latest release (version 24.0). The most important change is that we now use HMMER3, the latest version of the popular profile hidden Markov model package. This software is approximately 100 times faster than HMMER2 and is more sensitive due to the routine use of the forward algorithm. The move to HMMER3 has necessitated numerous changes to Pfam that are described in detail. Pfam release 24.0 contains 11,912 families, of which a large number have been significantly updated during the past two years. Pfam is available via servers in the UK (http://pfam.sanger.ac.uk/), the USA (http://pfam.janelia.org/) and Sweden (http://pfam.sbc.su.se/).
14,075 citations
TL;DR: Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems are indicated.
Abstract: A 2.91-billion base pair (bp) consensus sequence of the euchromatic portion of the human genome was generated by the whole-genome shotgun sequencing method. The 14.8-billion bp DNA sequence was generated over 9 months from 27,271,853 high-quality sequence reads (5.11-fold coverage of the genome) from both ends of plasmid clones made from the DNA of five individuals. Two assembly strategies-a whole-genome assembly and a regional chromosome assembly-were used, each combining sequence data from Celera and the publicly funded genome effort. The public data were shredded into 550-bp segments to create a 2.9-fold coverage of those genome regions that had been sequenced, without including biases inherent in the cloning and assembly procedure used by the publicly funded group. This brought the effective coverage in the assemblies to eightfold, reducing the number and size of gaps in the final assembly over what would be obtained with 5.11-fold coverage. The two assembly strategies yielded very similar results that largely agree with independent mapping data. The assemblies effectively cover the euchromatic regions of the human chromosomes. More than 90% of the genome is in scaffold assemblies of 100,000 bp or more, and 25% of the genome is in scaffolds of 10 million bp or larger. Analysis of the genome sequence revealed 26,588 protein-encoding transcripts for which there was strong corroborating evidence and an additional approximately 12,000 computationally derived genes with mouse matches or other weak supporting evidence. Although gene-dense clusters are obvious, almost half the genes are dispersed in low G+C sequence separated by large tracts of apparently noncoding sequence. Only 1.1% of the genome is spanned by exons, whereas 24% is in introns, with 75% of the genome being intergenic DNA. Duplications of segmental blocks, ranging in size up to chromosomal lengths, are abundant throughout the genome and reveal a complex evolutionary history. Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems. DNA sequence comparisons between the consensus sequence and publicly funded genome data provided locations of 2.1 million single-nucleotide polymorphisms (SNPs). A random pair of human haploid genomes differed at a rate of 1 bp per 1250 on average, but there was marked heterogeneity in the level of polymorphism across the genome. Less than 1% of all SNPs resulted in variation in proteins, but the task of determining which SNPs have functional consequences remains an open challenge.
12,098 citations
TL;DR: In a four-genome analysis of 3' UTRs, approximately 13,000 regulatory relationships were detected above the estimate of false-positive predictions, thereby implicating as miRNA targets more than 5300 human genes, which represented 30% of the gene set.
11,624 citations
TL;DR: A mature web tool for rapid and reliable display of any requested portion of the genome at any scale, together with several dozen aligned annotation tracks, is provided at http://genome.ucsc.edu.
Abstract: As vertebrate genome sequences near completion and research refocuses to their analysis, the issue of effective genome annotation display becomes critical. A mature web tool for rapid and reliable display of any requested portion of the genome at any scale, together with several dozen aligned annotation tracks, is provided at http://genome.ucsc.edu. This browser displays assembly contigs and gaps, mRNA and expressed sequence tag alignments, multiple gene predictions, cross-species homologies, single nucleotide polymorphisms, sequence-tagged sites, radiation hybrid data, transposon repeats, and more as a stack of coregistered tracks. Text and sequence-based searches provide quick and precise access to any region of specific interest. Secondary links from individual features lead to sequence details and supplementary off-site databases. One-half of the annotation tracks are computed at the University of California, Santa Cruz from publicly available sequence data; collaborators worldwide provide the rest. Users can stably add their own custom tracks to the browser for educational or research purposes. The conceptual and technical framework of the browser, its underlying MYSQL database, and overall use are described. The web site currently serves over 50,000 pages per day to over 3000 different users.
9,605 citations
TL;DR: Velvet represents a new approach to assembly that can leverage very short reads in combination with read pairs to produce useful assemblies and is in close agreement with simulated results without read-pair information.
Abstract: We have developed a new set of algorithms, collectively called "Velvet," to manipulate de Bruijn graphs for genomic sequence assembly. A de Bruijn graph is a compact representation based on short words (k-mers) that is ideal for high coverage, very short read (25-50 bp) data sets. Applying Velvet to very short reads and paired-ends information only, one can produce contigs of significant length, up to 50-kb N50 length in simulations of prokaryotic data and 3-kb N50 on simulated mammalian BACs. When applied to real Solexa data sets without read pairs, Velvet generated contigs of approximately 8 kb in a prokaryote and 2 kb in a mammalian BAC, in close agreement with our simulated results without read-pair information. Velvet represents a new approach to assembly that can leverage very short reads in combination with read pairs to produce useful assemblies.
9,389 citations
References
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TL;DR: Evidence is presented suggesting that chromosomal rearrangements that determine the lengths of these segments are randomly distributed within the genome.
Abstract: Linkage relationships of homologous loci in man and mouse were used to estimate the mean length of autosomal segments conserved during evolution. Comparison of the locations of greater than 83 homologous loci revealed 13 conserved segments. Map distances between the outermost markers of these 13 segments are known for the mouse and range from 1 to 24 centimorgans. Methods were developed for using this sample of conserved segments to estimate the mean length of all conserved autosomal segments in the genome. This mean length was estimated to be 8.1 +/- 1.6 centimorgans. Evidence is presented suggesting that chromosomal rearrangements that determine the lengths of these segments are randomly distributed within the genome. The estimated mean length of conserved segments was used to predict the probability that certain loci, such as peptidase-3 and renin, are linked in man given that homologous loci are chi centimorgans apart in the mouse. The mean length of conserved segments was also used to estimate the number of chromosomal rearrangements that have disrupted linkage since divergence of man and mouse. This estimate was shown to be 178 +/- 39 rearrangements.
602 citations
TL;DR: It is suggested that increased monoamine oxidase A activity is a risk factor for panic disorder in female patients because the longer alleles (3a, 4 and 5) were more active than allele 3.
Abstract: A genetic contribution to the pathogenesis of panic disorder has been demonstrated by clinical genetic studies. Molecular genetic studies have focused on candidate genes suggested by the molecular mechanisms implied in the action of drugs utilized for therapy or in challenge tests. One class of drugs effective in the treatment of panic disorder is represented by monoamine oxidase A inhibitors. Therefore, the monoamine oxidase A gene on chromosome X is a prime candidate gene. In the present study we investigated a novel repeat polymorphism in the promoter of the monoamine oxidase A gene for association with panic disorder in two independent samples (German sample, n = 80; Italian sample, n = 129). Two alleles (3 and 4 repeats) were most common and constituted >97% of the observed alleles. Functional characterization in a luciferase assay demonstrated that the longer alleles (3a, 4 and 5) were more active than allele 3. Among females of both the German and the Italian samples of panic disorder patients (combined, n = 209) the longer alleles (3a, 4 and 5) were significantly more frequent than among females of the corresponding control samples (combined, n = 190, chi2 = 10.27, df = 1, P = 0.001). Together with the observation that inhibition of monoamine oxidase A is clinically effective in the treatment of panic disorder these findings suggest that increased monoamine oxidase A activity is a risk factor for panic disorder in female patients.
597 citations
TL;DR: It is proposed that the differences arising because mutation patterns vary with the timing of replication of different chromosomal regions in the germline can account for both the origin of isochores in mammalian genomes and the observation that silent nucleotide substitutions in different mammalian genes do not have the same molecular clock.
Abstract: In the traditional view of molecular evolution, the rate of point mutation is uniform over the genome of an organism and variation in the rate of nucleotide substitution among DNA regions reflects differential selective constraints1,2. Here we provide evidence for significant variation in mutation rate among regions in the mammalian genome. We show first that substitutions at silent (degenerate) sites in protein-coding genes in mammals seem to be effectively neutral (or nearly so) as they do not occur significantly less frequently than substitutions in pseudogenes. We then show that the rate of silent substitution varies among genes and is correlated with the base composition of genes and their flanking DNA. This implies that the variation in both silent substitution rate and base composition3 can be attributed to systematic differences in the rate and pattern of mutation over regions of the genome. We propose that the differences arise because mutation patterns vary with the timing of replication of different chromosomal regions in the germline. This hypothesis can account for both the origin of isochores in mammalian genomes4 and the observation5 that silent nucleotide substitutions in different mammalian genes do not have the same molecular clock.
594 citations
TL;DR: The emerging concept is that only three mechanisms of amplification are responsible for the vast majority of interspersed repeats and that with each autonomous element a number of dependent non-autonomous sequences have co-amplified.
588 citations
TL;DR: Several characteristics of EST-verified splice sites are analyzed and weight matrices for the major groups are built, which can be incorporated into gene prediction programs and should be significant for future investigations of the splicing mechanism.
Abstract: A set of 43 337 splice junction pairs was extracted from mammalian GenBank annotated genes. Expressed sequence tag (EST) sequences support 22 489 of them. Of these, 98.71% contain canonical dinucleotides GT and AG for donor and acceptor sites, respectively; 0.56% hold non-canonical GC-AG splice site pairs; and the remaining 0.73% occurs in a lot of small groups (with a maximum size of 0.05%). Studying these groups we observe that many of them contain splicing dinucleotides shifted from the annotated splice junction by one position. After close examination of such cases we present a new classification consisting of only eight observed types of splice site pairs (out of 256 a priori possible combinations). EST alignments allow us to verify the exonic part of the splice sites, but many non-canonical cases may be due to intron sequencing errors. This idea is given substantial support when we compare the sequences of human genes having non-canonical splice sites deposited in GenBank by high throughput genome sequencing projects (HTG). A high proportion (156 out of 171) of the human non-canonical and EST-supported splice site sequences had a clear match in the human HTG. They can be classified after corrections as: 79 GC-AG pairs (of which one was an error that corrected to GC-AG), 61 errors that were corrected to GT-AG canonical pairs, six AT-AC pairs (of which two were errors that corrected to AT-AC), one case was produced from non-existent intron, seven cases were found in HTG that were deposited to GenBank and finally there were only two cases left of supported non-canonical splice sites. If we assume that approximately the same situation is true for the whole set of annotated mammalian non-canonical splice sites, then the 99.24% of splice site pairs should be GT-AG, 0.69% GC-AG, 0.05% AT-AC and finally only 0.02% could consist of other types of non-canonical splice sites. We analyze several characteristics of EST-verified splice sites and build weight matrices for the major groups, which can be incorporated into gene prediction programs. We also present a set of EST-verified canonical splice sites larger by two orders of magnitude than the current one (22 199 entries versus approximately 600) and finally, a set of 290 EST-supported non-canonical splice sites. Both sets should be significant for future investigations of the splicing mechanism.
581 citations